The field of industrial microbiology comprises the testing of environments, equipment and manufacturing facilities for sterility. The principal means by which such sterility testing is conducted is by obtaining an appropriate sample from the environment of concern and conducting a microbiological assay with the use of appropriate microbiological test media. The most basic problem encountered in the field of industrial microbiology is that of assuring that the test medium that is used will not itself contaminate the environment to be tested. Various degrees of sterility assurance level (SAL) are known. For example, in the United States the most commonly encountered SAL is 10xe2x88x923, meaning that the level of contamination of vessels containing test media is xe2x89xa61 in 1000 vessels that test positive for microbial contamination, while the international SAL is 10xe2x88x926. For ultrasensitive sterility testing of clean rooms, surgical environments, pharmaceutical manufacturing environments and the like, an ultrapure test medium having an SAL of 10xe2x88x926 is most desirable, yet difficult to achieve with standard methods.
Two common methods of achieving an SAL of 10xe2x88x926 are by autoclaving and by irradiation with gamma rays. However, the drawback to autoclaving is that the outside of the container quickly becomes nonsterile following autoclaving upon exposure to a nonsterile environment. The drawback to gamma irradiation with respect to glass and plastic containers is that gamma rays cause clear glass to yellow and plastic to darken, making any growth in the media difficult to detect.
The present invention overcomes these drawbacks of the prior art by imparting an SAL of 10xe2x88x926 to test media that is relatively long lasting and that may be achieved simply and efficiently. The essence of the invention is the exposure of fluid test media in plastic containers to a beam of electrons.
The generation of high energy electrons has recently become commercially available. High energy electrons are a form of ionizing radiation capable of killing microorganisms by creating highly reactive free radicals that break down the DNA of microorganisms, rendering them incapable of reproduction. At the same time, such ionizing radiation may attack the polymers of plastic vessels, causing depolymerization, grafting, crosslinking and chain scission and so care must be taken to select plastic vessels formed from polymers that are not susceptible to such unwanted chemical reactions. Known commercial sources of electron beam irradiation include Iotron Technologies, Inc. of British Columbia Canada and Titan Scan Systems of Denver, Colo. Iotron Technologies offers a 60 KWatt/10 MeV electron beam accelerator having a dosing range of from 0 to 100 kGy per pass.
According to the present invention, it has been discovered that test media having a typical bioburden on the order of xe2x89xa625 colony-forming units (CFU) may be sterilized to an SAL of 10xe2x88x926 by exposing the same to electron beam irradiation at a dosage of from about 15 kGy to about 30 kGy at a rate of from about 175 to about 225 cm/min., preferably 18 to 25 kGy at about 200 cm/min.
In an especially preferred embodiment, the test media vessels are bottles formed from cyclohexanedimethanol-modified polyethylene wherein the cap assembly of such a bottle is formed from polyethylene and polypropylene and the bottle optionally has a septum in the cap assembly formed from butyl rubber. Following irradiation by an electron beam no undesirable chemical reactions involving the polymer of the vessel and no undesirable darkening or yellowing is observed. A preferred arrangement of such sterility media-containing bottles for exposure to the electron beam irradition is 6 packs of 20 bottles each arranged uniformly on an exposure tray measuring approximately 40 inchesxc3x9750 inches; each bottle has a 200 mL volume that is filled with approximately 100 mL fluid test media and each bottle is laid on its side so that the depth of the media is xe2x89xa61xc2xd inches. Average bioburden may be ascertained in accordance with the ANSI/AAMI/ISO 11137 protocol, Method 1, this bioburden number being ascertained prior to exposure of the media to the electron beam irradiation. The ANSI/AAMI/ISO 11137 Dose Audit protocol is preferably used for validation and periodic audits of the SAL.